PhiX:Plaque assay

Summary

This protocol is usually used in conjunction with serial dilution of a stock solution of phage for enumeration of phage titer. But it can also be used with any phage-containing input. With thanks to the Wichman lab for setting me up with a good protocol for this early on.

Media Preparation
The usual media for plating is lysogeny broth (LB). Remember that this is not a defined media (so you should attempt to minimise batch effects for reproducibility) and it comes in Lennox and Miller varieties. The appropriate type is Miller which contains 10g/L of sodium chloride (versus 5g/L in Lennox). Both plate (base layer) and top agars should be prepared. For one litre: 10 g Tryptone 5 g yeast extract 10 g NaCl 15g agar (if plate agar) | 7g (if top agar)

Make up to 1 litre in distilled water and autoclave. After autoclaving:
CaCl₂ to a final concentration of 2 mM, MgCl₂ to a final concentration of 10 mM. (for these use hydrated crystals as anhydrous powders do not dissolve effectively).

Plating Procedure
Add 100ul

Recommended Reading
I recommend checking:
Kropinski, A.M., Mazzocco, A., Waddell, T.E., Lingohr, E. and Johnson, R.P., 2009. Enumeration of bacteriophages by double agar overlay plaque assay. Bacteriophages: Methods and Protocols, Volume 1: Isolation, Characterization, and Interactions, pp.69-76.
for more details of the plaque assay and variations thereon.